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Identification of peroxisome proliferator-activated receptor alpha (PPARα)-dependent genes involved in peroxisome proliferator-induced short-term pleiotropic responses using fluorescent differential display technique.

Lee Wing Sum. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 206-226). / Abstracts in English and Chinese. / Abstract --- p.i / Abstract (Chinese Version) --- p.iv / Acknowledgements --- p.vii / Table of Contents --- p.viii / List of Abbreviations --- p.xiv / List of Figures --- p.xvii / List of Tables --- p.xxiv / Chapter Chapter 1 --- Introduction --- p.1 / Chapter Chapter 2 --- Literature review --- p.3 / Chapter 2.1 --- Peroxisomes --- p.3 / Chapter 2.2 --- Peroxisome proliferators --- p.5 / Chapter 2.3 --- Human exposure pathways to peroxisome proliferators --- p.5 / Chapter 2.4 --- Peroxisome proliferator-induced pleiotropic effects in rodents --- p.7 / Chapter 2.4.1 --- Short-term effects --- p.7 / Chapter 2.4.1.1 --- Hepatomegaly --- p.7 / Chapter 2.4.2.1 --- Peroxisome proliferation --- p.8 / Chapter 2.4.1.3 --- Alteration of gene transcriptions --- p.8 / Chapter 2.4.2 --- Long-term effect --- p.9 / Chapter 2.5 --- Mechanisms of actions of peroxisome proliferators --- p.9 / Chapter 2.5.1 --- Substrate overload --- p.9 / Chapter 2.5.2 --- Receptor-mediated --- p.11 / Chapter 2.6 --- Peroxisome proliferator-activated receptors (PPARs) --- p.11 / Chapter 2.6.1 --- Structure of PPARs --- p.11 / Chapter 2.6.2 --- Tissue-specific expression of PPARs --- p.15 / Chapter 2.6.3 --- Physiological functions of PPARs --- p.19 / Chapter 2.6.3.1 --- PPARα --- p.19 / Chapter 2.6.3.2 --- PPARγ --- p.21 / Chapter 2.6.3.3 --- PPARδ --- p.23 / Chapter 2.7 --- Role of PPARα involved in peroxisome proliferator-induced pleiotropic responses --- p.24 / Chapter 2.7.1 --- Short-term effects --- p.24 / Chapter 2.7.2 --- Long-term effect --- p.24 / Chapter 2.8 --- Mechanisms of peroxisome proliferator-induced hepatocarcinogenesis --- p.25 / Chapter 2.8.1 --- Oxidative stress --- p.25 / Chapter 2.8.2 --- Suppression of apoptosis --- p.26 / Chapter 2.8.3 --- Increased cell proliferation --- p.27 / Chapter 2.9 --- Species difference to peroxisome proliferator-induced pleiotropic effects --- p.28 / Chapter 2.10 --- Fluorescent differential display (FDD) --- p.32 / Chapter Chapter 3 --- Objectives --- p.35 / Chapter Chapter 4 --- Materials and methods --- p.37 / Chapter 4.1 --- Animals and treatments --- p.37 / Chapter 4.1.1 --- Materials --- p.37 / Chapter 4.1.2 --- Methods --- p.37 / Chapter 4.2 --- Serum triglyceride and cholesterol analyses --- p.39 / Chapter 4.2.1 --- Materials --- p.41 / Chapter 4.2.2 --- Methods --- p.41 / Chapter 4.2.2.1 --- Serum preparation --- p.41 / Chapter 4.2.2.2 --- Triglyceride determination --- p.41 / Chapter 4.2.2.3 --- Cholesterol determination --- p.42 / Chapter 4.3 --- Statistical analysis --- p.42 / Chapter 4.4 --- Tail-genotyping --- p.42 / Chapter 4.4.1 --- Materials --- p.44 / Chapter 4.4.2 --- Methods. --- p.44 / Chapter 4.4.2.1 --- Preparation of genomic tail DNA --- p.44 / Chapter 4.4.2.2 --- PCR reaction --- p.45 / Chapter 4.5 --- Total RNA isolation --- p.45 / Chapter 4.5.1 --- Materials --- p.48 / Chapter 4.5.2 --- Methods --- p.48 / Chapter 4.6 --- DNase I treatment --- p.48 / Chapter 4.6.1 --- Materials --- p.49 / Chapter 4.6.2 --- Methods --- p.49 / Chapter 4.7 --- Reverse transcription of mRNA and fluorescent PCR amplification --- p.50 / Chapter 4.7.1 --- Materials --- p.50 / Chapter 4.7.2 --- Methods --- p.53 / Chapter 4.8 --- Fluorescent differential display (FDD) --- p.53 / Chapter 4.8.1 --- Materials --- p.53 / Chapter 4.8.2 --- Methods --- p.54 / Chapter 4.9 --- Excision of differentially expressed cDNA fragments --- p.54 / Chapter 4.9.1 --- Materials --- p.57 / Chapter 4.9.2 --- Methods --- p.57 / Chapter 4.10 --- Reamplification of differentially expressed fragments --- p.57 / Chapter 4.10.1 --- Materials --- p.60 / Chapter 4.10.2 --- Methods --- p.60 / Chapter 4.11 --- Subcloning of reamplified cDNA fragments --- p.62 / Chapter 4.11.1 --- PCR-TRAP® cloning system --- p.62 / Chapter 4.11.1.1 --- Materials --- p.63 / Chapter 4.11.1.2 --- Methods --- p.63 / Chapter 4.11.2 --- AdvaTage´ёØ PCR cloning system --- p.65 / Chapter 4.11.2.1 --- Materials --- p.65 / Chapter 4.11.2.2 --- Methods --- p.66 / Chapter 4.12 --- Purification of plasmid DNA from recombinant clones --- p.69 / Chapter 4.12.1 --- Materials --- p.69 / Chapter 4.12.2 --- Methods --- p.69 / Chapter 4.13 --- DNA sequencing of differentially expressed cDNA fragments --- p.70 / Chapter 4.13.1 --- CEQ 2000 Dye Terminator Cycle Sequence system --- p.71 / Chapter 4.13.1.1 --- Materials --- p.71 / Chapter 4.13.1.2 --- Methods --- p.71 / Chapter 4.13.2 --- ABI PRISM´ёØ dRhodamine Terminator Cycle Sequencing system --- p.72 / Chapter 4.13.2.1 --- Materials --- p.72 / Chapter 4.13.2.2 --- Methods --- p.72 / Chapter 4.13.3 --- Homology search against computer databases --- p.73 / Chapter 4.14 --- Northern analysis of differentially expressed cDNA fragments --- p.73 / Chapter 4.14.1 --- Formaldehyde gel electrophoresis of total RNA --- p.74 / Chapter 4.14.1.1 --- Materials --- p.74 / Chapter 4.14.1.2 --- Methods --- p.74 / Chapter 4.14.2 --- Preparation of cDNA probes for hybridization --- p.74 / Chapter 4.14.2.1 --- PCR DIG labeling --- p.75 / Chapter 4.14.2.1.1 --- Materials --- p.75 / Chapter 4.14.2.1.2 --- Methods --- p.75 / Chapter 4.14.2.2 --- Random Prime cDNA DIG labeling --- p.75 / Chapter 4.14.2.2.1 --- Materials --- p.75 / Chapter 4.14.2.2.2 --- Methods --- p.76 / Chapter 4.14.3 --- Purification of DNA from agarose gel --- p.77 / Chapter 4.14.3.1 --- Materials --- p.77 / Chapter 4.14.3.2 --- Methods --- p.78 / Chapter 4.14.4 --- Hybridization --- p.78 / Chapter 4.14.4.1 --- Materials --- p.78 / Chapter 4.14.4.2 --- Methods --- p.73 / Chapter 4.14.5 --- Synthesis of mouse GAPDH probe from normalization --- p.80 / Chapter 4.14.5.1 --- Materials --- p.80 / Chapter 4.14.5.2 --- Methods --- p.80 / Chapter Chapter 5 --- Results --- p.82 / Chapter 5.1 --- Liver morphology --- p.82 / Chapter 5.2 --- Liver weight --- p.82 / Chapter 5.3 --- Serum triglyceride and cholesterol levels --- p.88 / Chapter 5.4 --- Confirmation of genotypes --- p.91 / Chapter 5.5 --- DNase I treatment --- p.91 / Chapter 5.6 --- FDD RT-PCR and band excision --- p.98 / Chapter 5.7 --- Reamplification of excised cDNA fragments --- p.111 / Chapter 5.8 --- Subcloning of reamplified cDNA fragments --- p.121 / Chapter 5.9 --- DNA sequencing of subcloned cDNA fragments --- p.124 / Chapter 5.10 --- Confirmation of the differentially expressed cDNA fragments by Northern blot analysis --- p.132 / Chapter 5.11 --- Temporal expression pattern of differentially expressed genes --- p.157 / Chapter 5.12 --- Tissue distribution pattern of differentially expressed genes --- p.171 / Chapter Chapter 6 --- Discussions --- p.183 / Chapter 6.1 --- "Lack of hepatomegaly, hypotriglyceridemia and hepatic nodule formation in PPARα (-/-) mice" --- p.184 / Chapter 6.2 --- "Identification of PPARα-dependent and Wy-14,643 responsive genes" --- p.185 / Chapter 6.3 --- Functional roles of the isolated cDNA fragments --- p.186 / Chapter 6.3.1 --- Fragments B14 and H4 --- p.187 / Chapter 6.3.2 --- Fragment H1 --- p.189 / Chapter 6.3.3 --- Fragment H5 --- p.192 / Chapter 6.3.4 --- Fragment H8 --- p.194 / Chapter 6.4 --- Temporal expression patterns of the isolated cDNA fragments --- p.196 / Chapter 6.5 --- Tissue distribution patterns of the isolated cDNA fragments --- p.197 / Chapter Chapter 7 --- Conclusions --- p.200 / Chapter Chapter 8 --- Future studies --- p.204 / Chapter 8.1 --- Subcloning and characterization of the other differentially expressed genes --- p.204 / Chapter 8.2 --- Overexpression and inhibition expression of specific genes --- p.204 / Chapter 8.3 --- Generating transgenic mice with target disruption of specific gene --- p.205 / References --- p.206

Identiferoai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_323194
Date January 2000
ContributorsLee, Wing Sum., Chinese University of Hong Kong Graduate School. Division of Biochemistry.
Source SetsThe Chinese University of Hong Kong
LanguageEnglish, Chinese
Detected LanguageEnglish
TypeText, bibliography
Formatprint, xxv, 226 leaves : ill. (some col., some mounted) ; 30 cm.
RightsUse of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/)

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